PROJECT SUMMARY Burkitt lymphoma (BL) is the most common childhood cancer in sub-Saharan Africa, outnumbering all other forms of pediatric tumors combined. BL is lethal if left untreated; however, the current treatment (intensive chemotherapy) is only 50% effective and difficult to implement in sub-Saharan Africa. To date, the vast majority of African cases of BL are associated with Epstein-Barr virus (EBV) infection. Thus, there is a critical need for a safe and effective vaccine to prevent and treat EBV and its associated diseases. The objective of the proposed study is to develop, characterize, and validate gp42-gH/gL-EBNA1-LMP2 virus-like particles (VLPs) in pre-clinical studies as a candidate vaccine against EBV. Previous approaches to EBV vaccine development have been limited due in part to the oncogenic potential of the virus genome and lack of animal models to test vaccine candidates. This proposal will examine a new strategy to develop a safe and effective vaccine against EBV, using VLPs that can present properly folded EBV proteins. To target both arms of the immune system?antibody-mediated and T-cell mediated?the vaccine will incorporate multiple EBV surface and intracellular antigens. The EBV protein complex gp42-gH/gL and tumor-associated Epstein- Barr nuclear antigen 1 (EBNA1) and latent membrane protein (LMP2) are prime candidates for use in EBV vaccine development. There is strong evidence that antibodies to the EBV protein complex gp42-gH/gL can neutralize EBV infection. EBNA1 and LMP2 are consistently expressed in B cells of all EBV+ BL patients and are recognized by CD4+ and CD8+ T cells. Thus, we hypothesize that gp42-gH/gL-EBNA1-LMP2 VLPs will generate robust EBV-neutralizing antibody responses and EBV-specific T-cell responses in vitro and in a humanized mouse model. The VLPs will be generated, characterized, and validated using wild-type mice (Aim 1), an in vitro model of BL (Aim 2), and humanized mice (Aim 2) A polyvalent vaccine that induces both prophylactic neutralizing antibodies and elicits therapeutic human T-cell responses will not only be an invaluable candidate vaccine in preventing EBV infection, but also of utmost importance in preventing and/or treating EBV-associated disease. If the vaccine is immunogenic, following completion of the proposal, we will utilize the current good manufacturing practice facility available at City of Hope to mass-produce clinical grade gp42-gH/gL-EBNA1-LMP2 VLPs and begin a phase 1 clinical study in 10-15 healthy adults to establish safety profiles and determine the protective efficacy of the candidate vaccine. In the long term, if successful, our approach will introduce a new prophylactic/therapeutic vaccine to the market with a potential for preventing or treating over 200,000 annual cancer cases associated with EBV, including 24,000 cases of BL in African children.